Process for the preparation of biotin

ABSTRACT

A novel process is described for preparation of biotin comprising preparation of substituted 3H, 5H-imidazo[1, 5c]tetrahydro thiazoles by contacting the boron trifluoride adduct of an appropriate thiazoline with the metallic derivative of an ester enolate, reducing the ester, hydrolyzing the thiazolidine moiety and hydrolyzing or oxidizing the resultant compound. Intermediates obtained in the preparation of biotin by the above process and alternate procedures for preparing said intermediates are also presented. A novel process for preparation of d-biotin is also given.

This is a division of application Ser. No. 379,247, filed on May 17,1982, now U.S. Pat. No. 4,468,516.

Biotin is a water-soluble vitamin required by higher animals and by manymicroorganisms. Biosynthesis of biotin by selected yeasts, molds andbacteria is well known. U.S. Pat. No. 3,393,129 reports the use of ad-biotin-producing strain of bacteria of the genus Sporobolomyces forcommercial production of this vitamin. Chemical synthesis is reported inU.S. Pat. Nos. 2,489,235; 2,489,236; 4,029,647 and 4,124,595.

As industrial demand for d-biotin increases, the search for improvedsynthetic processes continues.

The present invention relates to a novel process for the preparation ofbiotin and novel intermediates useful therein.

One class of intermediates of the present invention are novel compoundsof formula II ##STR1## wherein

X is sulfur or oxygen;

R₁ is --(CH₂)₄ CH₃, or --(CH₂)₃ OR or --(CH₂)₅ OR wherein R is alkyl, or--(CH₂)₄ CN, or --(CH₂)₄ COOR' wherein R' is alkyl or phenyl;

R₂ and R₃ when taken together are cycloalkyl or --CH₂ --CH₂ --Y--CH₂--CH₂ wherein Y is sulfur, oxygen or NR" wherein R" is COOR'" whereinR'" is alkyl, or

R₂ and R₃ when taken separately, are each alkyl, cycloalkyl or phenyl,provided that R₂ and R₃ are not both phenyl;

R₄ is hydrogen, alkyl, alkoxyalkyl, cycloalkyl, monoalkyl, substitutedcycloalkyl, phenyl or mono-, di or trialkyl substituted phenyl; and

when R₄ is hydrogen, the addition salts thereof;

said alkyl and alkoxy having from 1 to 4 carbon atoms and saidcycloalkyl having from 5 to 7 carbon atoms.

Preferred compounds include those wherein R₁ is --(CH₂)₄ CH₃ or --(CH₂)₄COOR' wherein R' is alkyl; R₂ and R₃ when taken together are cycloalkyl,or R₂ and R₃ when taken separately are each alkyl; and R₄ is alkyl.Especially preferred of these compounds are those wherein R₁ is --(CH₂)₄CH₃ or --(CH₂)₄ COOCH₃ ; R₂ and R₃ when taken together are cyclohexyl,or R₂ and R₃ taken separately are each methyl; and R₄ is ethyl.

Also within the scope of the present invention are intermediates usefulfor the preparation of thiazoles of formula II. Thus, the presentinvention includes compounds of formula I ##STR2## wherein

R₁, R₂ and R₃ are as previously defined;

R₂ and R₃ when taken together are cycloalkyl or --CH₂ --CH₂ --Y--CH₂--CH₂ wherein Y is sulfur, oxygen or NR" wherein R" is COOR'" whereinR'" is alkyl; or

R₂ and R₃ when taken separately are each alkyl, cycloalkyl or phenyl,provided that R₂ and R₃ are not both phenyl;

said alkyl having from 1 to 4 carbon atoms and said cycloalkyl havingfrom 3 to 7 carbon atoms.

Also included in the present invention are the boron trifluoride adductsof compounds of formula I, especially those wherein R₁ is --(CH₂)₄ CH₃or --(CH₂)₄ COOR' wherein R' is alkyl; and

R₂ and R₃ when taken together are cycloalkyl, or

R₂ and R₃ when taken separately are each alkyl.

Preferred compounds include those wherein R₁ is --(CH₂)₄ CH₃ or --(CH₂)₄COOCH₃ ; and

R₂ and R₃ when taken together are cyclohexyl, or

R₂ and R₃ when taken separately are each methyl.

The present invention also includes a process for the preparation ofcompounds of formula II comprising contacting a boron trifluoride adductof a compound of formula I with a compound of the formula

    MiR.sub.4 O.sub.2 C--CH--A]

wherein

A is --N═C═O, --N═C═S, --N═S═O, --N--CO₂ R₇, or --N═C(H)R₇ wherein R₇ isalkyl or phenyl;

M is a metal selected from lithium, sodium, potassium, zinc, magnesiumor zirconium or a counterion of the formula N(R₈)₄ ⁺ or B(R₈)₂ whereinR₈ is an alkyl group having from 1 to 4 carbon atoms; and

R₄ is as previously defined.

A further class of intermediates useful in the process of the presentinvention are novel compounds of formula III ##STR3## wherein X, R₁, R₂and R₃ are as previously defined.

Preferably R₁ is --(CH₂)₄ CH₃ or --(CH₂)₄ COOR' wherein R' is alkyl; R₂and R₃ when taken together are cycloalkyl, or R₂ and R₃ when takenseparately are each alkyl. Especially preferred are compounds wherein R₁is --(CH₂)₄ CH₃ or --(CH₂)₄ COOCH₃ ; and

R₂ and R₃ when taken together are cyclohexyl or

R₂ and R₃ when taken separately are each methyl.

Further intermediates of the present invention are novel compounds offormula IV ##STR4## wherein

X, R₁, R₂ and R₃ are as previously defined; and

R₅ is --C(O)R₆ or --SO₂ R₆ wherein R₆ is alkyl, haloalkyl, phenyl ormono- or dialkyl substituted phenyl or camphoryl; said alkyl having from1 to 4 carbon atoms and said cycloalkyl having from 5 to 7 carbon atoms.

Preferred compounds include those wherein

R₁ is --(CH₂)₄ CH₃ or --(CH₂)₄ COOR' wherein R' is alkyl;

R₂ and R₃ when taken together are cycloalkyl, or

R₂ and R₃ when taken separately are each alkyl; and

R₅ is acetyl, mesyl, tosyl or camphorsulfonyl.

Especially preferred are compounds wherein

R₁ is --(CH₂)₄ CH₃ or --(CH₂)₄ COOCH₃ ; and

R₂ and R₃ when taken together are cyclohexane or

R₂ and R₃ when taken separately are each methyl; and

R₅ is --SO₂ R₆ wherein R₆ is d-10-camphoryl.

Further intermediates of the present invention are novel compounds offormula V ##STR5## wherein X and R₁ are as previously defined and R₅ isas previously defined or hydrogen.

Preferred compounds include those wherein R₁ is --(CH₂)₄ CH₃ or --(CH₂)₄COOCH₃ ; and

R₅ is hydrogen, acetyl, mesyl, tosyl or camphorsulfonyl. Especiallypreferred are compounds wherein R₁ is --(CH₂)₄ CH₃ and R₅ isd-10-camphorsulfonyl or hydrogen.

Further intermediates of the present invention are those of formula VI##STR6## wherein X and R₁ are as previously defined, provided that whenR₁ is (CH₂)₄ COOR', R' is alkyl having from 2 to 4 carbon atoms.

Also included in the present invention are processes for preparation ofcompounds of Formula VI by cyclizing intermediate compounds of formulaV.

Further intermediates of the present invention are compounds of theformula ##STR7## wherein X and R₁ are as previously defined and Y is ═O,formula VIII, or Y is --H and --OH, formula IX.

Compounds VI may also be prepared by reducing a thiolactone of formulaVIII in a reaction inert solvent with an akali metal borohydridefollowed by treatment with an electropositive metal in the presence ofan acid.

Compounds VI wherein X is sulfur may be converted to the oxygen analogby contacting it with a haloalcohol in the presence of weak base in aprotic solvent.

The present invention further comprises a method for preparation ofbiotin comprising contacting in solution a compound of formula IIA withan alkali metal borohydride followed by the addition of water, treatingthe resultant compound of formula III with strong aqueous acid or withan alkyl or aryl sulfonyl halide or acyl halide in the presence of base,e.g. trimethylamine, triethylamine or pyridine and contacting theseproducts with strong aqueous acid, and, when X is sulfur, refluxing witha haloalcohol. The resultant compound may be hydrolyzed, treated withacid followed by sodium diethyl malonate and then hydrolyzed, oroxidized depending on the nature of the R₁ group to form biotin. Thuswhen R₁ is --(CH₂)₄ CH₃, the resultant product is oxidized; when R₁ is--(CH₂)₄ COOR', the resultant product is hydrolyzed; when R₁ is --(CH₂)₄CN, the resultant product is hydrolyzed; when R₁ is --(CH₂)₅ OR, theresultant product is hydrolyzed and oxidized; and when R₁ is --(CH₂)₃ ORthe resultant product is treated with acetic acid saturated withhydrogen bromide followed by sodium diethyl malonate, the resultantdiester is hydrolyzed with barium hydroxide, and the mixture heated at atemperature of about 180° until reaction is substantially complete.

In a preferred process of this invention, d-biotin may be prepared byresolving an acid of formula IIA with (d)-ephedrine, separating theresultant diastereometric mixture of compounds, esterifying therequisite stereoisomer to give a compound of formula II wherein R₄ ismethyl, contacting this ester with borohydride and then acid, and when Xis sulfur, treating the resultant bicyclic thiourea with a haloalcohol,and hydrolyzing or oxidizing, as appropriate in view of the nature ofthe R₁ group, to form biotin.

The present invention relates to the synthesis of biotin from theintermediates described above as shown in Schemes A and B to whichreference is made for the following discussion. The formulae given inthese Schemes and throughout the present application conform to theaccepted convention for indicating stereoisomers, namely " " to indicatean atom projecting into the plane of the paper (α-orientation) " " toindicate an atom projecting out from the plane of the paper(β-orientation) and hence the plane of the molecule itself and "˜" toindicate a substituent which is in either the α or β-orientation.Numbering of compounds throughout this application follows the sequencegiven in Schemes A and B. It will be appreciated that R₂, R₃ and R₄groups in the compounds of formulae I to V are protecting groups whichwill be subsequently removed in later reaction steps. Likewise R₅, R₆,R, R', R" and R'" are intermediate groups in the synthesis of the finalcompounds. Accordingly, while intermediates having R₂, R₃, R₄, R₅, R₆,R, R', R" and R'" substituents as previously defined are preferred foruse in the present invention, the use of such substituents is notcritical and other similar protecting and intermediate substituentsgroups may be employed in the present process to obtain biotin. Forexample, higher alkyl or cycloalkyl groups, up to about 17 carbon atoms,may be employed, together with substituted aryl groups, such as phenylsubstituted with alkyl, halo, nitro or alkoxy groups, or naphthyl.

Substituted 3H,5H-imidazo[1,5c]tetrahydrothiazoles of formula II may beprepared by contacting in a nonprotic solvent at about -100° C. to -30°C. the boron trifluoride adduct of a compound of formula I, wherein R₁,R₂ and R₃ are as previously defined, with the metallic derivatives of anester enolate of the formula ##STR8## wherein M⁺ is lithium, sodium,potassium, zinc, magnesium or zirconium or a counterion of the formulaN(R₈)₄ ⁺ or B(R₈)₂ wherein R₈ is an alkyl group having from 1 to 4carbon atoms and A is --N═C═O or --N═C═S. Compounds wherein A is asdefined are preferred for use in the present invention, but it will beappreciated that similar protecting groups may be employed to make theintermediates useful for the preparation of biotin by the process of thepresent invention, for example compounds wherein A is --N═S═O, --N--COORwherein R is alkyl or --N═C(H)R₇ wherein R₇ is phenyl.

More particularly, compounds of formula II wherein R₁ is --(CH₂)₄ CH₃ or--(CH₂)₄ COOR' wherein R' is alkyl, R₂ and R₃ when taken together arecycloalkyl or --CH₂ --CH₂ --N(R")--CH₂ --CH₂ -- wherein R" is --COOR'"wherein R'" is alkyl or R₂ and R₃ separately are each alkyl, cycloalkylor phenyl provided that R₂ and R₃ are not both phenyl; and

R₄ is alkyl or phenyl may be prepared by contacting in a non-proticsolvent, preferably tetrahydrofuran, the boron trifluoride adduct of acompound of formula I wherein R₁ is --(CH₂)₄ CH₃ or --(CH₂)₄ CO₂ R'wherein R' is alkyl and R₂ and R₃ are as given above, at a temperaturebetween about -100° C. to -0° C. preferably near -78° C. with a metalloderivative of an ester enolate of the formula ##STR9## wherein

[M] is lithium, sodium, potassium, zinc or magnesium, but mostpreferably lithium;

A, when X of the resultant compound of formula II is oxygen, is --N═C═Oor A, when X of said compound of formula II is sulfur, is --N═C═S; and

R₄ is alkyl or phenyl.

Preferred compounds which may be prepared by this method include thosewherein R₁ is --(CH₂)₄ CH₃ or --(CH₂)₄ COOCH₃, R₂ and R₃ when takentogether are cycloalkyl, preferably cyclohexyl or --CH₂ --CH₂ --Y--CH₂--CH₂ -- wherein Y is NR" wherein R" is --COOR'" wherein R'" is alkyl,preferably methyl or ethyl, R₂ and R₃ when taken separately are alkyl,preferably methyl or ethyl or R₂ and R₃ when taken together arecycloalkyl, preferably cyclohexyl and R₄ is alkyl, preferably ethyl,n-propyl or isopropyl, alkoxy, preferably ethylmethoxy or alkylsubstituted phenyl, preferably 2,6-di-t-butyl-4-methyl phenyl or2-methyl-6-t-butylphenyl or cycloalkyl such as norboronyl.

The boron trifluoride adduct of the compound of formula I may beprepared by combining in a suitable non-polar solvent, preferablytetrahydrofuran, said thiazoline of formula I and an essentiallyequimolar amount of boron trifluoride diethyl ether at a temperaturebetween about -78° C. and 30° C. preferably about 0° C.

The metallo derivative of ester enolate may be prepared by standardmethods such as combining in a suitable non-polar solvent, preferablytetrahydrofuran at temperatures between about -100° C. and 10° C.preferably -78° C. an alkyl isocyanatoacetate and a metallodialkylamide, for example lithium diisopropylamide, which in turn isgenerated by adding butyllithium to a dialkylamine solution. In the caseof an alkyl or phenyl isothiocyanatoacetate, either a metallodialkylamide preferably lithium di-isopropylamide or a metallo alkoxide,preferably lithium t-butoxide can be used to generate the metalloderivatives of the ester enolates. The nature of the alkyl or arylsubstituent (R₄) present in the isothiocyanatoacetate influences theratio of products of formulas IIA:IIB formed. For example, the ratio ofIIA:IIB is 1.3:1 when methyl isothiocyanatoacetate is used and withethyl isothiocyanatoacetate, the ratio of IIA:IIB is 3:1. Since IIA isthe desired isomer in the synthesis of biotin, compounds in which R₄ isethyl are preferred.

The desired compound of formula IIA can be separated from formula IIB bystandard chromatographic processes or by crystallization. The compoundsof formula IIA are employed in the further synthesis of biotin.

3-Thiazolines of formula I may, in turn, be prepared according to themethod of Thiel, Asinger and Schmiedel (Liebigs Ann. Chem. 611 121(1958)) wherein 2-bromoaldehydes, compounds easily synthesized by knownmethods, of the formula ##STR10## wherein R₁ is as defined, is combinedwith sodium hydrogen sulfide and then with a carbonyl compound of theformula ##STR11## wherein R₂ and R₃ are as defined, followed by theaddition of ammonia.

Compounds of formula I, for example, which may be prepared by thismethod, are those wherein R₁ is preferably --(CH₂)₄ CH₃ or --(CH₂)₄COOCH₃ ; and R₂ and R₃ together are preferably cyclohexyl or R₂ and R₃separately are each methyl.

The desired enantiomer of the compound of formula IIA required for asynthesis of d-biotin can be obtained by a resolution of racemicmixtures of IIA in which R₄ =H. The overall procedure may beaccomplished, for example, by a saponification of the racemic ester IIAobtained in the imine addition reaction to form a racemic acid whichwhen treated with a chiral base, may be separated into diasteriometricsalts, which once separated may be converted to optically pure esters offormula IIA. More specifically, racemic ester IIA (R₄ =CH₂ CH₃) may besaponified to the corresponding racemic acid of formula IIA (R₄ =H) upontreatment with an alkali hydroxide such as sodium hydroxide in a polarsolvent such as methanol or tetrahydrofuran. Treatment of racemic acidof formula IIA with an optically pure base such as d-ephedrine in apolar solvent such as ether generates a solid which can be crystallizedin an optically pure form. Treatment of this salt, for example, with analcoholic solvent in the presence of acid such as methanolic hydrogenchloride generates optically pure ester of formula IIA wherein R₄ ismethyl.

Compounds of formula III,7-hydroxymethyl-3H,5H-imidazo[1,5c]tetrahydrothiazoles, may be preparedby contacting esters of formula IIA wherein R₄ is preferably ethyl ormethyl in a polar solvent, preferably methanol, ethanol ortetrahydrofuran with a borohydride derivative, an alkali metalborohydride for example, wherein the alkali metal is preferably sodium,at a temperature between -10° C. and 25° C. for a period of about 1-5hours followed by addition of water.

Alternatively, optically pure acids of formula IIA may be reduced withdiborane to generate III directly.

Compounds of Formula II may then be converted directly to the biotinring structure of formula VI via intermediate V. For example, alcoholIII is treated with a strong acid at elevated temperatures to generateVI directly. In particular, compounds of formula VI wherein R₁ is--(CH₂)₄ CH₃ or --(CH₂)₄ CO₂ H and X is O or S may be prepated bycontacting alcohol III, wherein R₁ is --(CH₂)₄ CH₃ or --(CH₂)₄ CO₂ R'wherein R' is preferably methyl, R₂ and R₃ when taken together arecyclohexyl, or when taken separately are alkyl preferably methyl withaqueous trifluoroacetic acid or methanesulfonic acid at temperaturebetween about 40° to 105° C. until reaction is substantially complete.

Alternatively, compounds of formula III may be converted to compounds offormula IV by contacting compounds of formula III with a sulfonyl oracyl halide in a polar solvent in the presence of a base, preferablytrialkylamine.

Thus compounds of formula IV wherein R₅ is SO₂ R₆ or COR₆ wherein R₆ isalkyl or haloalkyl, most preferably methyl, tolyl or camphoryl areprepared by contacting III wherein R₁ is --(CH₂)₄ CH₃ or --(CH₂)₄ CO₂R', R₂ and R₃ when taken together are cyclohexyl or when takenseparately are alkyl, preferably methyl, in a solvent such as methylenechloride with triethylamine and an appropriate sulfonyl chloride attemperatures between about -78° to 25° C. When racemic III is treatedwith an optically active sulfonyl chloride, a diastereomeric mixtureresults which may be separated to afford optically pure compounds offormula IV of the desired chirality. For example, compound IV wherein R₅is SO₂ R₆ and R₆ is d- or l-10-camphoryl and R₁ is (CH₂)₄ CH₃ and R₂ andR₃ when taken together are cyclohexyl can easily be separated by meansof silica gel or alumina chromatography to give the desired purediastereomer of formula IV.

Compounds of formula IV may then be converted directly to the biotinring structure by treating compounds of formula IV wherein R₅ is acyl orsulfonyl in a strong acid at elevated temperatures. For example, thed-biotin framework of VI in which R₁ is (CH₂)₄ CH₃ may be generated bytreating the requisite camphorsulfonate of formula IV in which R₁ is(CH₂)₄ CH₃, R₂ and R₃ taken together are cyclohexyl, R₅ is SO₂ R₆ and R₆is d-10-camphor with aqueous trifluoroacetic acid at temperature betweenabout 35° to 105° C. for 1 to 24 hours.

When R₁ is --(CH₂)₄ COOR', the resultant product is hydrolyzed; when R₁is --(CH₂)₄ CN, the resultant product is hydrolyzed; when R₁ is --(CH₂)₅OR, the resultant product is hydrolyzed and oxidized; and when R₁ is--(CH₂)₃ OR the resultant product is treated with acetic acid saturatedwith hydrogen bromide followed by sodium diethyl malonate, the resultantdiester is hydrolyzed with barium hydroxide, and the mixture heated at atemperature between 150° to 200° until reaction is substantiallycomplete.

Compounds of formula VI wherein X is S may be converted to the oxygenanalogue by contacting the corresponding thiourea derivative of formulaVI wherein X is sulfur with a haloalcohol, preferably bromoethanol, inan polar solvent such as ethanol, methoxyethanol or diglyme, andrefluxing under inert gas, preferably nitrogen, until reaction isessentially complete, from 2 to 24 hours, and then treating with a weakbase, an alkali metal carbonate for example, preferably a saturatedsolution of sodium carbonate.

The conversion of compound VI wherein X is O and R₁ is (CH₂)₄ CH₃ tobiotin may be accomplished by a microbiological oxidation. The preferredmicrobiological oxidation is that disclosed in Ogino et al in U.S. Pat.No. 3,859,167, the disclosure of which is incorporated herein byreference. Accordingly, biotin wherein R₁ is (CH₂)₄ CO₂ H is obtainedupon treatment of VI wherein R₁ is (CH₂)₄ CH₃ and X is oxygen with theorganism Corynebacterium primorioxydans. Compound VI wherein X is S andR₁ is --(CH₂)₄ CH₃ may likewise be converted by microbiologicaloxidation by an organism such as Corynebacterium primorioxydans to thesulfur analog of biotin.

A novel process is also herein presented for preparation of intermediatecompounds of formula VI by reduction of the corresponding thiolactone offormula VIII A as shown in Scheme B wherein R₁, R₂, R₃, R₄ and X are aspreviously defined.

As shown in Scheme B, thiolactones of formula VIIIA wherein R₁ is (CH₃)₄CH₃ may be prepared by:

contacting a compound of the formula IIA wherein R₁ is --(CH₂)₄ CH₃, R₂and R₃ are each alkyl, preferably methyl, and R₄ is alkyl, preferablyethyl, with a strong acid, preferably aqueous trifluoroacetic acid at atemperature between 80° to 120° C., preferably about 100° C.; or

contacting a compound of the formula IIA in a polar solvent, forexample, aqueous methanol, with an essentially equimolar amount of base,preferably an alkali metal hydroxide, for a period of about 5 to 12hours at a temperature between 20° C. to 35° C. followed byacidification to a pH between 2.0 to 3.0, preferably about 2.5 with anaqueous acid halide, preferably hydrochloric acid; and

contacting the resultant carboxylic acid with acid, preferablytrifluoroacetic acid with an excess molar amount of water at atemperature between 45° C. to 55° C. for about 6 to 8 hours.

Alternatively, compounds of formula VIII A may be prepared by:

contacting IIB in a polar solvent, for example, aqueous methanol, withan essentially equimolar amount of base, preferably an alkali metalhydroxide, for a period of 1 to 2 hours at a temperature of -0°-10° C.followed by acidification to a pH between 2.0 to 3.0, preferably about2.5 with an aqueous acid halide, preferably hydrochloric acid;

contacting the resultant acid with acid, preferably trifluoroacetic acidin an excess molar amount of water at a temperature of about 15° C.-35°C. for 2 to 3 hours;

contacting the resultant thiol VIIB in a polar solvent, preferablymethylene chloride, with a basic trialkyl-amine, preferablytriethylamine followed by an alkylhaloformate, preferably ethylchloroformate at a temperature between about 15° C. to 35° C. for aperiod of 2 to 3 hours to give lactone VIIIB which may be converted toVIIIA by contacting VIIB in a polar solvent, preferably tetrahydrofuran,with a non-nucleophalic base, for example.

Intermediate compounds of formula VIIIA may be converted to intermediatecompounds of formula VI by reduction. Compounds of formula VI, forexample, wherein R₁ is preferably --(CH₂)₄ CH₃ or --(CH₂)₄ COOCH₃ may beprepared by contacting a thiolactone of formula VIIA wherein R₁ is--(CH₂)₄ CH₃ or --(CH₂)₄ COOCH₃ in a polar solvent, preferably methanolat a temperature between -10° C. to 25° C., preferably about 0° C. witha metallic borohydride, preferably sodium borohydride for a period ofabout one hour and contacting the resultant hemiacetal in acid solutionwith zinc metal under reflux for a period of 12 to 48 hours, untilreduction is essentially complete. ##STR12##

The present invention is illustrated by the following examples. Itshould be understood, however, that the invention is not limited to thespecific details of these examples.

EXAMPLE 1 3H, 5H-Imidazo[1, 5c]thiazole-7-carboxylic acid,tetrahydro-3,3-dimethyl-5-oxo-1-pentyl-, ethyl ester (1α,7α,7aα) and(1α, 7β,7aα)

To a tetrahydrofuran solution (50 ml ) containing2,2-dimethyl-5-pentyl-3-thiazoline (5.58 grams, 30.2 mmole) at 0° C. wasadded boron trifluoride etherate (3.70 ml, 30.2 mmol) over a one minuteperiod. Solution was allowed to warm to room temperature and stirred forone hour and then cooled to -78° C. Diisopropylamide was prepared byadding 2.3M m-butyllithium (13.1 ml, 30.2 mmol) to diisopropylamine(4.24 ml, 30.2 mmol) in tetrahydrofuran (300 ml) at -78° C. and stirringfor 60 minutes. To this solution was added drop-wise over a period of 1minute, ethylisocyanoacetate (3.90 grams, 30.2 mmol). This solution wasstirred at -78° C. for 5 minutes and then was added over a 1 minuteperiod to the boron trifluoridethiazoline solution. The mixture wasstirred at -78° C. for two hours, allowed to warm gradually to roomtemperature and stirred for another 1 hour. The reaction mixture wasconcentrated. Ethyl acetate was added and the organic solution wasextracted with 0.5N HCL, dried over magnesium sulfate and concentratedin vacuo. The crude product was purified by column chromatography onsilica gel (eluant methylene chloride:ether, 3:2) to give 4.69 g (50%)of a product mixture (1:1) containing 3H,5H-imidazo[1,5c]thiazole-7-carboxylic acid,tetrahydro-3,3-dimethyl-5-oxo-1-pentyl ethylester (1α,7α,7aα) MP71°-73°.

IR (KBr) 3267, 2926, 1731, 1704; NMR (d, CDCL₃) 0.6-2.4 (20H, m, CH₃,CH₂) 3.2-3.7 (1H, m, CHS), 3.9-4.6 (4H, m, CHN, CHN, OCH₂), 5.1-5.4 (1H,m, NH). Analysis Calculated for C₁₅ H₂₆ O₃ N₂ S: C, 57.32; H, 8.28, N,8.92. Found: C, 56.97; H, 8.12; N, 8.87; And3H,5H-Imidazo[1,5c]thiazole-7-carboxylic acid,tetrahydro-3,3-dimethyl-5-oxo-1-pentyl-, ethylester (1α,7β,7aα), MP74°-75° C. IR(KBr) 3280, 3926, 1731, 1705; NMR (d, CDCL₃) 0.66-2.25(20H, m, CH₃ CH₂) 3.0-3.5 (1H, m, CHS), 3.9-4.6 (4H, m, CHN, CHN, OCH₂),5.3-5.6 (1H, m, NH) Analysis Calculated for C₁₅ H₂₆ O₃ N₂ S: C, 57.32;H, 8.28; N, 8.92; S, 10.19; Found: C, 57.47; H, 8.28; N, 8.97; S, 10.18.

EXAMPLE 2 3H,5H-Imidazo[1,5c]thiazole-7-carboxylic acid,tetrahydro-3,3-dimethyl-5-thioxo-1-pentyl-, ethyl ester (1α,7α,7aα) and(1α,7β,7aα)

2,2-Dimethyl-5-pentyl-3-thiazoline (860 mg, 4.65 mmol) was dissolved intetrahydrofuran (20 ml) and cooled to 0° C. Boron trifluoride etherate(0.510 ml, 4.65 mmol) was added over a one minute period. The solutionwas allowed to warm to room temperature for about 1.25 hour and thencooled to -78° C. In a separate flask was placed diisopropylamine (0.652ml, 4.65 mmol) followed by tetrahydrofuran (10 ml). The solution wascooled to -78° C. 1.5M butyl lithium (3.1 ml, 4.65 mmol) was added overa 5 minute period. The solution was stirred at -78° for one hour. Tothis solution was added ethyl isothiocyanotoacetate (674 mg, 4.65 mol)in tetrahydrofuran (5 ml) over a 5 minute period. The solution wasstirred for 25 minutes at -78° and was then added to the borontrifluoride thiazoline solution. The solution was stirred at -78° for 2hours and quenched with acetic acid (266 ml, 4.66 mmol). The solutionwas allowed to warm to room temperature, concentrated in vacuo and wastaken up in methylene chloride. This organic solution was washed withaqueous bicarbonate, dried over MgSO₄ and concentrated to afford a blackoil which was purified by column chromatography on pH 9 silica gel(eluant methylene chloride:ether, 20:1) to give 1.030 g (67%) of aproduct mixture containing 278 mg (18%) of3H,5H-imidazo[1,5c]thiazole-7-carboxylic acid,tetrahydro-3,3-dimethyl-5-thioxo-1-pentyl; ethylester (1α,7α,7aα) whichformed needles after a methanol recrystallization,

mp 123-124.5. IR(KBr) 3207, 2933, 1743; NMR (d, CDCL₃) 0.6-2.4(20H, m,CH₂, CH₃), 3.2-3.8 (1H, m, CHS), 4.0-4.8 4H,M,CHN, CH₂ --O), 6.8-7.0(1H, m, NH). Analysis Calculated for C₁₅ H₂₆ O₂ N₂ S₂ : C, 54.51: H,7.93; N, 8.48. Found C, 54.44; H 7.80; N 8.62; and 752 mg (49%) of3H,5H-imidazo[1,5c]thiazole-7-carboxylic acid,tetrahydro-3,3-dimethyl-5-thioxo-1-pentyl; ethylester (1α,7β,7aα) whichwas crystallized from ether, MP 106°-107° C. IR (KBr) 3437, 2925, 1743;NMR (d, CDCl₃) 0.6-2.3 (20H, m, CH₂, CH₃), 3.0-3.6 (1H, m, CHS). 4.0-4.9(4H, m, CHN, CH₂ O), 6.3-6.5 (1H, m, NH). Analysis Calculated for C₁₅H₂₆ O₂ N₂ S_(2:) C, 54.51; H, 7.93; N, 4.48. Found C, 54.23; H, 7.71; N,8.63.

EXAMPLE 3 3H,5H-Imidazo[1,5c]thiazole-1-pentanoic acid,tetrahydro-7-carboethoxy-3,3-dimethyl-5-oxo, methyl ester, (1α,7α,7aα)and (1α,7β,7aα)

To 2,2-Dimethyl-3-thiazole-5-pentanoic acid, methyl ester (5.77 g, 23mmol) in dry tetrahydrofuran (50 ml) at 0° C. was added over a oneminute period boron trifluoride etherate (2.82 ml, 23 mmol). Thereaction mixture was allowed to warm to room temperature, stirred forone hour, and cooled to -78° C. To an addition funnel containingdiisopropylamine (3.23 ml, 23 mmol) at room temperature intetrahydrofuran (150 ml) was added 2.3M butyl lithium (10 ml, 23 mmol).The solution was stirred at room temperature for 15 minutes and thencooled to -78° C. To this solution was added all at once ethylisocyanatoacetate (2.97 g, 23 mmol) in tetrahydrofuran (10 ml). Thissolution was allowed to stir for 6 minutes and then added to the borontrifluoride-thiazoline solution. The resulting solution was stirred at-78° for 2 hours and allowed to warm to room temperature and stirred forabout 20 minutes. The reaction mixture was concentrated, taken up inethyl acetate and extracted with 0.5N hydrochloric acid solution. Theaqueous layer was back extracted with ethyl acetate (3X). The organicswere washed with brine, dried over magnesium sulfate, concentrated toafford 9.8 grams of crude product which was chromatographed on 330 gramsof silica gel using methylene chloride: diethyl ether (3:2) to afford3.05 g (37%) of a product mixture (1:1) containing3H,5H-imidazo[1,5c]thiazole-1-pentanoic acid,tetrahydro-7-carboethoxy-3,3-dimethyl-5-oxo; methylester, (1α,7α,7aα) asan oil. IR(CHCl₃) 3444, 2926, 1726; NMR (d, CDCL₃) 1.1-2.1 (15H, m, CH₂,CH₃), 2.2-2.6 (2H, m, CHCH₂), 3.3-3.6 (1H, m, CHS), 3.7 (3H, s, OCH₃),4.0-4.6 (4H, m, CHN, OCH₂), 5.1-5.3 (1H, m, NH). Analysis Calculated forC₁₆ H₂₆ O₅ N₂ S: C, 53.61; H, 7.31; N, 7.81; S, 8.94. Found: C, 53.32;H, 7.26; N, 8.06; S, 8.57; and 3H,5H-Imidazo[1,5c]thiazole-1-pentanoicacid, tetrahydro-7-carboethoxy 3,3-dimethyl-5-oxo-methyl ester,(1α,7β,7aα),

mp 89°-90° C. IR(KBr) 3242, 2928, 1746, 1700; NMR (d, CDCL₃) 0.63-2.1(15H, m, CH₂, CH₃), 2.13-2.6 (2H, m, CHCH₂), 3.1-3.5 (1H, m, CHS) 3.7(3H, s, OCH₃), 3.9-4.6 (4H, m, CHN, OCH₂); 4.9-5.2 (1H, m, NH). AnalysisCalculated for C₁₆ H₂₆ O₅ N₂ S: C, 53.61; H, 7.31; N, 7.81; S, 8.94.Found: C, 53.81; H, 7.52; N, 7.75; S, 8.88.

EXAMPLE 4 3H,5H-Imidazo[1,5c]thiazole-1-pentanoic acid,tetrahydro-7-carboethoxy-3,3-dimethyl-5-thioxo, methyl ester (1α,7α,7aα)and (1α,7β,7aα)

To a dry tetrahydrofuran solution (100 ml) containing2,2-dimethyl-3-thiazoline-5-pentanoic acid, methyl ester (11.17 g, 48.8mmol) under a nitrogen atmosphere at -4° C. was added dropwise over a 2minute period boron trifluoride etherate (6.00 ml, 48.78 mmol). Internaltemperature did not rise abot 0° C. The reaction mixture was stirred at-4° to 0° C. for 15 minutes. The ice bath was removed and the reactionmixture was stirred for 45 minutes and then cooled to -75° C.

Lithium t-butoxide (4.10 grams, 51.22 mmol) was dissolved in drytetrahydrofuran (150 ml) and the solution was cooled to -75° C. Ethylisothiocyanatoacetate (7.07 grams, 48.78 mmol) was dissolved in drytetrahydrofuran (50 ml) in a cold jacketed addition funnel (-75° C.) andwas added to the lithium t-butoxide solution over 6 to 7 minutes.Internal temperature did not exceed -71° C. The solution was stirred for10 minutes following the addition. A polyethylene tube was put into theanion solution and nitrogen was used to push the anion into theimine/boron trifluoride solution. The addition occurred in less than oneminute. The internal temperature of the final reaction mixture rose from-75° to -65°. The reaction mixture was stirred at -75° C. for 1.5 hoursand then quenched with acetic acid (2.8 ml, 48.78 mmol) intetrahydrofuran (5 ml). The brown reaction mixture became light orange.Organic solvents were removed in vacuo and the residue was taken up in900 ml of ethyl acetate and washed with 5×200 ml of sodium bicarbonatesolution followed by 1×200 ml of brine. The organic portion was driedover magnesium sulfate, filtered and concentrated in vacuo to afford18.22 grams of product. The crude product was purified by columnchromatography on pH 9 buffered silica gel (eluant methylenechloride:ether, 98:2) to give 14.80 g (81%) of a product mixture (1:2.5)containing 3H,5H-imidazo[1,5c]thiazole-1-pentanoic acid,tetrahydro-7-carboethoxy-3,3-dimethyl-5-thioxo, methyl ester (1α,7α,7aα)which could be recrystallized from hexane to give a solid, mp 55°-55.5°C. IR(KBr) 3211, 2929, 1740; NMR (d, CDCL₃) 1.1-2.6 (17H, m, CH₂ CH₃,CH₂, CCH₃), 3.1-3.6 (1H, m, CHS), 3.7 (3H, S, OCH₃), 3.9-4.8 (4H, m,CHN, OCH₂), 6.5 (1H, m, NH). Analysis Calculated for C₁₆ H₂₆ N₂ O₄ S₂ :C, 51.34; H, 6.95; N, 7.49. Found C, 51.23; H, 6.86; N, 7.26; and3H,5H-imidazo[1,5c]thiazole-1-pentanoic acid,tetrahydro-7-carboethoxy-3,3-dimethyl-3-thioxo-methyl ester (1α,7β,7aα)which was recrystallized from hexane to give a solid, mp 76°-78° C.IR(KBr) 3439, 3411, 2940, 1740;

NMR (d, CDCl₃) 1.1-2.7 (17H, m, CH₂ CH₃, CH₂, CCH₃), 3.1-3.6 (1H, m,CHS), 3.7 (3H, s, OCH₃), 3.9-4.9 (4H, m, CHN, OCH₂), 5.9-6.3 (1H, m,NH). Analysis Calculated for C₁₆ H₂₆ N₂ O₄ S₂ : C, 51.34; N, 6.95; N,7.49. Found: C, 51.09; H, 6.88; N, 7.52.

EXAMPLE 5 3H,5H-Imidazo[1,5c]thiazole-7-carboxylic acid,tetrahydro-3,3-dimethyl-5-thioxo-1-pentyl,-,2,6-di-t-butyl-4-methylphenylester (1α,7α,7aα) and (1α,7β,7aα)

A procedure identical to that of Example 2 involving2,2-dimethyl-5-pentyl-3-thiazoline and 2,6-di-t-butyl.4-methylphenyl-2-isothiocyanatoacetate afforded a 1:5 mixture (90%yield) containing 3H,5H-imidazo[1,5c]thiazole-7-carboxylic acid,tetrahydro-3,3-dimethyl-5-thioxo-1-pentyl-2,6-di-t-butyl-4-methylphenylester (1α,7α,7aα), mp 87°-95° C. IR(KBr) 3189, 2958, 1762;

NMR (d, CDCL₃) 0.6-1.7 (29H, m, C(CH₃)₃, --CH₂, CH_(2--CH) ₃), 1.9 (3H,s, CCH₃), 2.2 (3H, s, CCH₃), 2.3 (3H, s, phenyl methyl), 3.2-3.8 (1H, m,CHS), 4.3-4.9 (1H, m, CHN), 6.9 (1H, bs, NH), 7.1 (2H, bs, Ar--H).Analysis calculated for C₂₈ H₄₄ N₂ O₂ S₂ : C, 66.62; H, 8.79; N, 5.55.Found: C, 66.60; H, 8.88; N, 5.52; and3H,5H-imidazo[1,5c]thiazole-7-carboxylic acid,tetrahydro-3,3-dimethyl-5-thioxo-1-pentyl-,2,6-di-t-butyl-4-methylphenylester (1α,7β,7aα), mp 149-151.

IR(KBr) 3447, 3177, 2958, 2924, 1760; NMR (d, CDCL₃) 0.57-2.2 (35H, m,C(CH₃)₃, --CH₂, --CH₂ --CH₃, C(CH₃)₂), 2.3 (3H, s, Ar--H), 3.6-4.2 (1H,m, CHS), 4.3-5.0 (1H, M, CHN), 6.3-6.6 (1H, m, NH), 7.1 (2H, bS, Ar--H).Analysis Calculated for C₂₈ H₄₄ N₂ O₂ S₂ : C, 66.62; H, 8.79; N, 5.55.Found: 66.56; H, 8.61; N, 5.60.

EXAMPLE 6 3H,5H-Imidazo[1,5c]thiazole-7-carboxylic acid,tetrahydro-3,3-pentamethylene-5-thioxo-1-pentyl, ethyl ester (1α,7β,7aα)

To a dry tetrahydrofuran solution (600 ml) containing2,2-pentamethylene-5-pentyl-3-thiazoline (123 g, 0.548 mol) under anitrogen atmosphere at -2° C. was added over a 10 minute period borontrifluoride etherate (67 ml, 0.548 mol). The reaction mixture wasstirred at 0° for 15 minutes. The ice bath was removed, the reactionmixture was stirred for 45 minutes and was then cooled to -78° C.Lithium t-butoxide (48.5 g, 0.603 mol) was dissolved in drytetrahydrofuran (800 ml) and the solution was cooled to -78° C. Ethylisothiocyanatoacetate (87.5 g, 0.603 mol) was dissolved in drytetrahydrofuran (250 ml) in a cold jacketed addition funnel (-78° C.)and was then added to the lithium t-butoxide solution over a 7 to 8minute period. The internal temperature did not exceed -68° C. Thesolution was stirred for an additional 17 minutes at which time apolyethylene tube was used with positive nitrogen pressure to push theanion and the imine/boron trifluoride (-78° C.) solution. The additionoccurred in about 3 minutes. The internal temperature rose from -78° to-55° C. The reaction mixture was stirred at -78° C. for 1.75 hour. andwas then quenched with acetic acid (36 ml, 0.603 mol) in tetrahydrofuran(40 ml). The reaction mixture was concentrated in vacuo and the residuewas taken up in ethyl acetate (3.25 liters) and washed with a 1:1mixture of aqueous brine and saturated sodium bicarbonate (2 liters)followed by aqueous brine (1 liter). The organic portion was dried overmagnesium sulfate, filtered and concentrated in vacuo to afford 212 g ofoily solids. A hexane:ether (12:1) trituration afforded 88.1 g of mainly3H,5H[1,5c]thiazole-7-carboxylic acid,tetrahydro-3,3-pentamethylene-5-thioxo-1-pentyl,-methyl ester(1α,7β,7aα). An additional 18.7 g was obtained by additional triturationof the mother liquor. An analytical sample, mp 121°-122° C. was obtainedafter carbon tetrachloride recrystallization. IR(KBr) 3429, 2930, 1741.NMR (d, CDCl₃) 0.8-2.2 (22H, m, CH₂, CH₃) 2.8-3.8 (3H, m, CHS, CCH₂),4.1-4.9 (4H, m, OCH₂, CHN), 6.5 (1H, bS, NH) Analysis Calculated for C₁₈H₃₀ N₂ O₂ S₂ : C, 58.38; H, 8.11; N, 7.57. Found C, 58.18; H, 7.98; N,7.74.

EXAMPLE 7 3H,5H-Imidazo[1,5c]thiazole-1-pentanoic acid,tetrahydro-7-carboethoxy-3,3-pentamethylene-5-thioxo, ethylester,(1α,7β,7aα)

To a dry tetrahydrofuran solution (40 ml) containing2,2-pentamethylene-3-thiazoline-5-pentanoic acid, methyl ester (10.12 g,37.6 mmol) under a nitrogen atmosphere at -10° C. was added over a 5minute period period boron trifluoride etherate (4.63 ml, 13.6 mmol).The reaction mixture was stirred at 0° for 15 minutes. The ice bath wasremoved, the reaction mixture was stirred for 45 minutes and was thencooled to -78° C. Lithium t-butoxide (3.31 g, 41.4 mmol) was dissolvedin dry tetrahydrofuran (50-75 ml) and the solution was cooled to -78° C.Ethy isothiocyanatoacetate (6.0 g, 41.4 mmol) was dissolved in drytetrahydrofuran (240 ml) in a cold jacketed addition funnel (-78° C.)and was then added to the lithium t-butoxide solution over a 10 minuteperiod. The internal temperature did not exceed -68° C. The solution wasstirred for an additional 15 minutes at which time a polyethylene tubewas used with positive nitrogen pressure to push the anion into theimine/boron trifluoride (-78° C.) solution. The addition including thewashes occurred in about 3 minutes. The reaction mixture was stirred at-78° C. for 1.75 hour and was then quenched with acetic acid (2.5 ml,41.4 mmol) in tetrahydrofuran (2 ml). The reaction mixture wasconcentrated in vacuo and the residue was taken up in ethyl acetate (250ml) and washed with a 1:1 mixture of aqueous brine and saturated sodiumbicarbonate (150 ml) followed by aqueous brine (75 ml). The organicportion was dried over magnesium sulfate, filtered and concentrated invacuo to afford 15.6 g of a reddish brown oil which was trituratedseveral times with a hexane:ether (10:1) solution to afford 8 g ofmainly 3H,5H-imidazo[1,5c]thiazole-1-pentanoic acid,tetrahydro-7-carboethoxy-3,3-pentamethylene-5-thioxo, ethyl ester(1α,7β,7aα). An analytical sample, mp 84°-87° was obtained from a carbontetrachloride recrystallization. IR(KBr) 3434, 2930, 1740, 1446, 1417NMR (d, CDCl₃) 1.05-2.2 (19H, m, CH₂, C--CH₃), 2.2-2.6 (2H, m, CH₂),3.2-3.6 (1H, m, CHS), 3.8 (3H, s, OCH₃), 4.2-5.0 (4H, m, CHN, OCH₂),6.2-6.35 (1H, bs, NH).

EXAMPLE 8 3H,5H-Imidazo[1,5c]thiazole-7-carboxylic acid,tetrahydro-3,3-dimethyl-5-thioxo-1-pentyl; methoxyethyl ester(1α,7α,7aα) and (1α,7β,7aα)

A procedure identical to that of Example 2 involving2,2-dimethyl-5-pentyl-3-thiazoline and methoxyethylisothiocyanatoacetate afforded after a pH 9 silica gel chromatography(eluant 98:2 methylene chloride:ether) a 1:2.9 mixture (85%) containing3H,5H-imidazo[1,5c]thiazole-7-carboxylic acid,tetrahydro-3,3-dimethyl-5-thioxo-1-pentyl; methoxyethyl ester(1α,7α,7aα), mp 69°-70° C. IR (KBr) 3437, 2926, 1756. NMR (d, CDCL₃)0.6-2.2 (17H, m, CH₂, CCH₃), 3.2-3.8 (6H, m, CHS, OCH₂, OCH₃) 4.1-4.8(4H, m, CHN, OCH₂), 6.6-6.8 (1H, bs, NH). Analysis Calculated for C₁₆H₂₈ N₂ S₂ O₃ : C, 53.30; H, 7.83; N, 7.77. Found: C, 53.18; H, 7.56; N,7.83; and 3H,5H-imidazo[1,5c]thiazole-7-carboxylic acid, tetrahydro-3,3-dimethyl-5-thioxo-1-pentyl, methoxyethyl ester (1α,7β,7aα), mp75°-77°. IR (KBr) 3437, 2926, 1756; NMR (d, CDCl₃) 0.6-2.4 (17H, m,CH₂,CCH₃) 3.0-3.8 (6H, m, CHS, OCH₂, OCH₃), 4.1-5.0 (4H, m, CHN, OCH₂),6.7-7.0 (1H, bS, NH). Analysis calculated for C₁₆ H₂₈ N₂ S₂ O₃ : C,53.30; H, 7.83; N, 7.77. Found: C, 53.08; H, 7.70; N, 7.87.

EXAMPLE 9 3H,5H-Imidazo[1,5c]thiazole-7-carboxylic acid,tetrahydro-3,3-dimethyl-5-thioxo-1-pentyl, n-propyl ester, (1α,7α,7aα)and (1α,7β,7aα).

A procedure identical to that of Example 2 involving2,2-dimethyl-5-pentyl-3-thiazoline and n-propyl isothiocyanatoacetateafforded after pH 9 silica gel chromatography (eluant 95:5 methylenechloride:ether) a 1:3 mixture (83% yield) containing3H,5H-imidazo[1,5c]thiazole-7-carboxylic acid,tetrahydro-3,3-dimethyl-5-thioxo-1-pentyl; n-propyl ester (1α,7α,7aα),mp 55°-56° C. IR (KBr) 3278, 2928, 1744; NMR (d, CDCL₃) 0.7-2.3 (22H, m,CH₂, CCH₃, CH₂ CH₃), 3.2-3.7 (1H, m, CHS), 4.0-4.8 (4H, m, CHN, OCH₂),6.5 (1H, bs, NH). Analysis calculated for C₁₆ H₂₈ N₂ S₂ O₂ : C, 55.78;H, 8.19, N, 8.13. Found C, 55.81; H, 8.03; N, 8.06. and3H,5H-imidazo[1,5c]thiazole-7-carboxylic acid, tetrahydro-3,3-dimethyl-5-thioxo-1-pentyl; n-propyl ester (1α,7β,7aα), mp 64°-66°C., IR (KBr) 3438, 2926, 1739; NMR (d, CDCL₃) 0.7-2.3 (22H, m, CH₂,CCH₃, CH₂ CH₃), 3.1-3.5 (1H, m CHS), 4.0-4.9 (4H, m, CHN, OCH₂), 6.6(1H, bs, NH) Analysis calculated for C₁₆ H₂₈ N₂ S₂ O₂ : C, 55.78; H,8.19; N, 8.13 Found C, 55.71; H, 7.96; N, 8.08.

EXAMPLE 10 3H,5H-Imidazo[1,5c]thiazole-7-carboxylic acid,tetrahydro-3,3-dimethyl-5-thioxo-1-pentyl; isopropyl ester, (1α,7α,7aα)and (1α,7β,7aα).

A procedure identical to that of Example 2 including2,2-dimethyl-5-pentyl-3-thiazoline and i-propyl-2-isothiocyanatoacetateafforded a 1:3 mixture (81% yield) containing3H,5H-imidazo[1,5c]thiazole-7-carboxylic acid,tetrahydro-3,3-dimethyl-5-thioxo-1-pentyl-, i-propyl ester (1α,7α,7aα),mp 103°-104° C. IR (KBr) 3210, 2956, 1737 NMR (d, CDCL₃) 0.6-2.5 (23H,m, CH₂, CH₃) 3.2-3.7 (1H, m, CHS), 4.0-5.4 (3H, m, CHN, CHO), 6.5-6.9(1H, bs, NH). Analysis Calculated for C₁₆ H₂₈ N₂ O₂ S₂ : C, 55.78; H,8.19; N, 8.13. Found: C, 56.01; 8.08; N, 8.15; and3H,5H-imidazo[1,5c]thiazole-7-carboxylic acid,tetrahydro-3,3-dimethyl-5-thioxo-1-pentyl, i-propyl ester (1α,7β,7aα),mp 46°-49° C. IR (KBr) 3245, 2977, 2557, 1737. NMR (d, CDCl₃) 0.7-2.3(23H, m, CH₂, CH₃), 3.1-3.6 (1H, m, CHS), 4.2 (1H, d, J=10 Hz, CHO),4.5-5.4 (2H, m, CHN), 6.7 (1H, bs, NH), Analysis Calculated for C₁₆ H₂₈N₂ O₂ S₂ : C, 55.78; H, 8.19; N, 8.13. Found: C, 55.61; H, 7.90; N,7.97.

EXAMPLE 11 3H,5H-Imidazo[1,5c]thiazole-7-carboxylic acid,tetrahydro-3,3-dimethyl-5-thioxo-1-pentyl, 2-methyl-6-t-butylphenylester (1α,7α,7aα) and (1α,7β,7aα).

A procedure identical to that of Example 2 involving2,2-dimethyl-5-pentyl-3-thiazoline and2-methyl-6-t-butylphenyl-2-isothiocyanatoacetate afforded a 1:3 mixture(90% yield) containing 3H,5H-imidazo[1,5c]thiazole-7-carboxylic acid,tetrahydro-3,3-dimethyl-5-thioxo-1-pentyl-, 2-methyl-6-t-butylphenylester (1α,7α,7aα), mp 136°-138° C. IR (KBr) 3193, 2928, 1753: NMR (d,CDCL₃) 0.6-1.7 (20H, m, CH₂ CH₂ CH₃, C(CH₃)₃) 1.9 (3H, s, C--CH₃), 2.13(3H, s, Ar--CH₃), 2.2 (3H, s, C--CH₃), 3.3-3.8 (1H, m, CHS), 4.4-5.0(2H, m, CHN), 6.9 (1H, bs, NH), 7.1-7.4 (3H, m, Ar--H), AnalysisCalculated for C₂₄ H₃₆ N₂ O₂ S₂ : C, 64.25; H, 8.09; N, 6.24. Found: C,64.27, H, 7.93, N, 6.41; and 3H,5H-imidazo[1,5c]thiazole-7-carboxylicacid, tetrahydro-3,3-dimethyl-5-thioxo-1-pentyl-,2-methyl-6-t-butylphenyl ester (1α,7β,7aα), mp 142°-144° C. IR (KBr)3217, 2925, 1745; NMR (d, CDCl₃) 0.7-1.8 (20H, m, CH₂, CH₂ CH₃,C(CH₃)₃), 1.95 (3H, s, C--CH₃), 2.1 (3H, s, C--CH₃) 2.15 (3H, s,Ar--CH₃), 3.6-4.2 (1H, m, CHS), 4.3-4.9 (2H, m, CHN), 6.5 (1H, bs, NH),7.0-7.4 (3H, m, Ar--H). Analysis Calculated for C₂₄ H₃₆ N₂ O₂ S₂ : C,64.25; H, 8.09; N, 6.24. Found: C, 64.04; H, 7.93; N, 6.10.

EXAMPLE 12 3H,5H-imidazo[1,5c]thiazole-7-carboxylic acid,tetrahydro-3,3-pentamethylene-5-thioxo-1-pentyl (1α,7β,7aα)

To a tetrahydrofuran solution (30 ml) containing3H,5H-imidazo[1,5c]thiazole-7-carboxylic acid,tetrahydro-3,3-pentamethylene-5-thioxo-1-pentyl, ethyl ester (1α,7β,7aα)(2.0 g 5.4 mmol) at 0° C. was added 2N sodium hydroxide (3.1 ml, 6.2mmol). The solution was stirred at 0° for 1 hour and allowed to stir atroom temperature for 17 hours. Acetic acid (355 ml, 6.21 mol) was thenadded, the solution was concentrated in vacuo and dissolved in ethylacetate (100 ml). The organic solution was extracted with 1N HCL (2×30ml) and the aqueous solutions were back washed with ethyl acetate. Thecombined organic extracts were dried over magnesium sulfate, filteredand concentrated to afford 1.83 g (100%) of3H,5H-imidazo[1,5c]thiazole-7-carboxylic acid,tetrahydro-3,3-pentamethylene-5-thioxo-1-pentyl (1α,7β,7aα),

NMR (d, CDCl₃ 0.62-2.3 (19H, m, CH₂, CH₃), 2.6-3.65 (3H, m, CHS, CCH₂),4.05-4.85 (2H, m, CHN), 7.3 (1H, bs, NH), 10.65 (1H, bs, OH).

EXAMPLE 13 3H,5H-imidazo[1,5c]thiazole-7-carboxylic acid,tetrahydro-3,3-pentamethylene-5-thioxo-1-pentyl, methyl ester IS(1α,7β,7aα)

To an ether solution (20 ml) containing3H,5H-imidazo[1,5c]thiazole-7-carboxylic acid,tetrahydro-3,3-pentamethylene-5-thioxo-1-pentyl (1α,7β,7aα) (761 mg,2.22 mol) at 0° C. was added 99% d-ephedrine (371 mg, 2.22 mol). Thesolution was stirred for 12 hours at 0° C. and filtered. The solids werewashed with ether, dried and recrystallized from benzene. Therecrystallized salt was then placed in a methanol solution (30 ml)saturated with hydrochloric acid (0° C.) and stirred for 3 hours. Theresultant solution was concentrated in vacuo, dissolved in ethyl acetate(50 ml) and washed with 2N hydrochloric acid (2×30 ml), dilute sodiumbicarbonate (1×30 ml) followed by brine (1×30 ml) and dried overmagnesium sulfate. The organic solution was filtered and concentrated invacuo to afford 346 mg (61%) of 3H,5H-imidazo[1,5c]thiazole-7-carboxylicacid, tetrahydro-3,3-pentamethylene-5-thioxo-1-pentyl, methyl ester IS(1α,7β,7aα)

NMR (d, CDCl₃) 0.7-2.4 (19H, m, CH₂, CH₃) 2.6-3.6 (3H, m, CHS, C-CH₂)3.78 (3H, S, OCH₃). 4.0-4.9 (2H, m, CHN), 6.85 (1H, bs, NH).

EXAMPLE 14 3H,5H-imidazo [1,5c]thiazole,tetrahydro-3,3-pentamethylene-7-hydroxymethyl-5-thioxo-1-pentyl 1S(1α,7β,7aα).

To a 1:1 tetrahydrofuran:methanol solution (8 ml) containing3H,5H-imidazo[1,5c]thiazole-7-carboxylic acid,tetrahydro-3,3-pentamethylene-5-thioxo-1-pentyl, methyl ester 1S(1α,7β,7aα) (657 mg, 1.85 mmol) was added sodium borohydride (274 mg,7.4 mol). The solution was stirred at 0° C. for 1.5 hour, at roomtemperature for 1.5 hour and concentrated under reduced pressure. Ethylacetate (75 ml) was added to the residue and the organic solution wasextracted with 0.5N HCL (1×40 ml) and a 1:1 brine: sodium bicarbonatesolution (1×40 ml). The organic layer was dried over magnesium sulfate;filtered and concentrated in vacuo to afford 580 mg (100%) of3H,5H-imidazo[1,5c]thiazole,tetrahydro-3,3-pentamethylene-7-hydroxymethyl-5-thioxo-1-pentyl IS(1α,7β,7aα).

NMR (d, CDCl₃) 0.6-2.2 (19H, m, CH₂, CH₃), 2.7-4.7 (7H, m, CH₂ --O, CHN,CHS, OH, C--CH₂), 7.05 (H, bs, NH).

EXAMPLE 15 3H,5H-Imidazo[1,5c]thiazole-1-pentanoic acid,tetrahydro-3,3-dimethyl-7-hydroxymethyl-5-thioxo, methyl ester,(1α,7β,7aα)

To a methanol solution (150 ml) containing3H,5H-imidazo[1,5c]thiazole-1-pentanoic acid,tetrahydro-7-carboethoxy-3,3-dimethyl-5-thioxo, methyl ester (1α,7β,7aα)(5.64 g, 15.08 mmol) at 0° C. was added sodium borohydride (2.28 g,60.32 mmol). The solution was stirred at 0° C. for 1.75 hour, andallowed to warm to room temperature (45 minutes). The reaction mixturewas concentrated in vacuo and taken up in ethyl acetate (300 ml). Theorganic solution was extracted with 0.2N HCL (70 ml) and a brinesolution (70 ml), dried over magnesium sulfate, filtered andconcentrated in vacuo to afford 5.0 g (99%) of3H,5H-imidazo[1,5c]thiazole-1-pentanoic acid,tetrahydro-3,3-dimethyl-7-hydroxymethyl-5-thioxo, methyl ester(1α,7β,7aα). An analytical sample, mp 103°-105°, was prepared by anether recrystallization. IR (KBr) 3411, 3199, 2927, 1731. NMR (d, CDCL₃)0.7-2.6 (14H, m, CH₂), 3.66 (3H, s, OCH₃), 2.9-4.9 (6H, m, CHN, CHS, CH₂--O, OH), 6.8-7.1 (1H, bs NH). Analysis Calculated for C₁₄ H₂₄ N₂ O₃ S₂: C, 50.60; H, 7.23; N, 8.43. Found: C, 50.36; H, 7.08; N, 8.49.

EXAMPLE 16 3H,5H-Imidazo[1,5c]thiazole,tetrahydro-3,3-dimethyl-7-hydroxymethyl-5-thioxo-1-pentyl, (1α, 7β, 7aα)

3H,5H-Imidazo[1,5c]thiazole-7-carboxylic acid,tetrahydro-3,3-dimethyl-5-thioxo-1-pentyl-, ethyl ester (1α,7β,7aα) (572mg, 1.73 mol) was dissolved in methanol (15 ml) and cooled to 0° C.Sodium borohydride (262 mg, 6.93 mol) was added and the solution wasstirred at 0° for 1.75 hr and allowed to come to room temperature. Thereaction mixture was concentrated in vacuo and taken up in ethyl acetate(75 ml). The organic solution was extracted with 0.5N HCL (1×40 ml) anda 1:1 brine:sodium bicarbonate solution (1×80 ml). The organic layer wasdried over magnesium sulfate, filtered and concentrated in vacuo toafford 486 mg (97%) of 3H,5H-Imidazo[1,5c]thiazole,tetrahydro-3,3-dimethyl-7-hydroxymethyl-5-thioxo-1-pentyl (1α,7β,7aα).An analytical sample, mp 112°-114° C. was obtained following a methanolrecrystallization. IR (KBr) 3351, 2925; NMR (d, CDCL₃) 0.4-2.5 (17H, m,C--CH₂, CH₃), 3.2-4.8 (6H, m, CHS, CHN, CH₂ O, OH), 6.9-7.2 (1H, m, NH)Analysis Calculated for C₁₃ H₂₄ N₂ OS₂ : C, 54.17; H, 8.33; N, 9.72.Found: C, 54.26; H, 8.07; N, 9.55.

EXAMPLE 17 3H,5H-Imidazo[1,5c]thiazole-1-pentanoic acid,tetrahydro-3,3-dimethyl-7-[[(methylsulfonyl)oxy]methyl]-5-oxo, methylester, (1α,7β,7aα)

To a methylene chloride solution (10 ml) containing3H,5H-imidazo[1,5c]thiazole-1-pentanoic acid,tetrahydro-3,3-dimethyl-7-hydroxymethyl-5-oxo, methyl ester (1α,7β,7aα).(220 mg, 0.695 mmol) at 0° C. was added triethylamine (200 μl, 1.42 mol,followed by methanesulfonyl chloride (621 μl, 0.775 mmol). The solutionwas allowed to warm to room temperature and stirred for 1 hour.Additional methylene chloride (50 ml) was added and the organic solutionwas extracted with H₂ O (1×30 ml), 0.5N HCl (1×20 ml), 5% sodiumbicarbonate (1×20 ml) and brine (1×20 ml) The aqueous layers werebackwashed with methylene chloride and the organic extracts were driedover magnesium sulfate, filtered and concentrated in vacuo to give 263mg (96%) of 3H,5H-imidazo[1,5c]thiazole-1-pentanoic acid,tetrahydro-3,3-dimethyl-7-[[(methylsulfonyl)oxy]methyl]-5-oxo, methylester (1α,7β,7aα). An analytical sample, mp 118.5°-119.5° C. wasobtained following an ether recrystallization. IR (KBr) 3305, 1732,1711. NMR (d, CDCL₃) 1.0-2.0 (12H, m, C--CH₂, C(CH₃)₂), 2.1-2.5##STR13## 3.1 ##STR14## 3.4-4.6 (8H, m, OCH₃, CH₂ --O, CHN, CHS),5.6-5.9 (1H, m, NH). Analysis Calculated for C₁₅ H₂₆ N₂ O₆ S₂ : C,45.68; H, 6.60; N, 7.11 Found: C, 45.90; H, 6.45; N, 7.08.

EXAMPLE 18 3H,5H-Imidazo[1,5c]thiazole,tetrahydro-3,3-pentamethylene-7-[[(d-camphorsulfonyl)oxy]methyl]-5-thioxo-1-pentyl1S (1α,7β,7aα)

To a methylene chloride solution (100 ml) containing3H,5H-imidazo[1,5c]thiazole,tetrahydro-3,3-pentamethylene-7-hydroxymethyl-5-thioxo-1-pentyl(1α,7β,7aα) (3.78 g, 11.5 mmol) at 0° C. was added triethylamine (1.61ml, 11.5 mol) followed by d-10-camphorsulfonyl chloride (2.89 g, 11.5mmol) in methylene chloride (25 ml). The reaction mixture was stirredfor 2 hr at 0° C. Additional methylene chloride (400 ml) was added andthe reaction mixture was washed with brine (1×100 ml), dried overmagnesium sulfate, filtered and concentrated in vacuo to afford a solidwhich was chromatographed on 500 g of 48-63μ silica gel usingether:methylene chloride (4:96). A total of 4.46 g (71%) ofdiasteriometric products was obtained. The first isomer (2.23 g) was 3H,5H imidazo [1,5c]thiazole,tetrahydro-3,3-pentamethylene-7-[[(d-camphorsulfonyl)oxy]methyl]-5-thioxo-1-pentyl 1S (1α,7β,7aα).sup.α D=+14.9 (C=0.01, methanol); NMR (d, CDCL₃)0.8 (3H, s, CCH₃), 1.0 (3H, s, CCH₃), 0.8-2.5 (28H, m, CH₂, C--CH, CH₂CH₃), 3.25 (2H, q CH₂ SO₂), 3.0-3.7 (1H, m, CHS), 3.7-4.7 (4H, m, CHN,CH₂ --O), 6.28 (1H, bs, NH).

EXAMPLE 19 Dl-Biotin, methyl ester

To a trifluoroacetic acid solution (10 ml) containing3H,5H-imidazo[1,5c]thiazole-1-pentanoic acid,tetrahydro-3,3-dimethyl-7-[[(methylsulfonyl)oxy]methyl]-5-oxo, methylester (1α,7β,7aα) (263 mg, 0.67 mmol) was added deuterium oxide (0.6ml). The solution was heated at 45° C. for 5 hours (reaction wasmonitored by NMR) and then concentrated in vacuo. The crude reactionmixture was dissolved in methylene chloride (200 ml) and extracted withdilute sodium bicarbonate (1×50 ml) followed by brine (1×50 ml). Theaqueous extracts were back extracted with methylene chloride (100 ml)and the combined orange extracts were dried over magnesium sulfate,filtered and concentrated under reduced pressure to give 183 mg of crudesolid which was triturated with ether and recrystallized from ethylacetate to give 78 mg (43%) or dl-biotin methyl ester, mp 127°-129° C.IR (KBr) 3225, 2941, 1751, 1718. NMR (d, DMSO) 1.15-1.95 (6H, m, CH₂),2.35 (2H, t, CH₂ --CO), 2.60-3.06 (2H, m, CH₂ S) 3.08-3.30 (1H, m, CHS),3.65 (3H, s, OCH₃), 4.15-4.64 (2H, m, CHN), 5.90 (1H, bs, NH), 6.13 (1H,bs, NH) Analysis Calculated for C₁₁ H₁₈ N₂ O₃ S: C, 51.16; H, 6.98; N,10.85. Found: C, 51.17; H, 7.01; N, 10.85. Mass Spectrum: Calculated(258.1038), observed (258.1041).

EXAMPLE 20 1-H-Thieno[3,4,-d]imidazole-4-pentanoic acid,hexahydro-2-thioxo (3aα,4β,6aα) (dl-thioxobiotin)

A trifluoroacetic acid solution (2 ml) containing3H,5H-imidazo[1,5c]thiazole,tetrahydro-3,3-pentamethylene-7-hydroxymethyl-5-thioxo-1-pentyl(1α,7β,7aα) (302 mg, 0.812 mmol) and water (0.86 ml) was heated underreflux for 1.5 hours and then cooled to 50° C. and concentrated invacuo. Ethanol (5 ml) was added and the solution was again concentratedunder reduced pressure. The solid residue was triturated withdiisopropylether and then ethyl acetate to afford 134 mg (64%) of1-H-thieno[3,4-d]imidazole-4-pentanoic acid, hexahydro-2-thioxo(3aα,4β,6aα) (dl-thioxobiotin) mp>250°. IR (KBr) 3407, 3291, 2939, 1694.NMR (d, DMSO) 1.22-1.77 (6H, m, CH₂), 2.33 (2H, t, J=8 HZ, CH₂ --CO),2.68 (1H, d, J=12 Hz, CH₂ S), 2.90 (1H, q, J_(AB) =12 HZ, Jax=6 HZ, CH₂S), 3.15-3.25 (1H, m, CHS), 4.35-4.50 (1H, m, CHN) 4.53-4.64 (1H, m,CHN), 8.21 (1H, bs, NH), 8.31 (1H, bs, NH). Analysis Calculated for C₁₀H₁₆ N₂ O₂ S₂ : C, 46.15; H, 6.15; N, 10.77, Found: C, 46.52; H, 6.19; N,10.48.

EXAMPLE 21 1-H-Thieno[3,4-d]imidazole-hexahydro-2-thioxo-4-pentyl; 3aS(3aα,4β,6aα)

3H,5Himidazo[1,5c]thiazole,tetrahydro-3,3-pentamethylene-7-[[(d-camphorsulfonyl)oxy]methyl]-5-thioxo-1-pentylIS(1α,7β,7aα) (761 mg, 1.40 mmol) was dissolved in trifluoroacetic acid(5 ml). Water (1 ml) was added and the solution was kept at 45° forabout 17 hours. The reaction mixture was concentrated in vacuo. Thewhite solid residue was dissolved in boiling ethyl acetate (500 ml),aqueous sodium bicarbonate (100 ml) was added and the hot two phasesystem was separated. The organic portion was dried over magnesiumsulfate, filtered and concentrated in vacuo to afford, after a methanoltrituration, 187 mg (58%) of1-H-thieno[3,4d]imidazolehexahydro-2-thioxo-4-pentyl 3aS (3aα,4β,6aα),mp 262°-262.5° C.; α_(D) =+133° (C=0.01, TFA); IR (KBr) 3220, 2919; NMR(d, DMSO) 0.68-1.04 (3H, m, CH₂ CH₃), 1.08-1.96 (8H, m, C--CH₂),2.64-2.98 (2H, m, CH₂ S), 3.05-3.36 (1H, m, CHS), 4.28-4.46 (1H, m,CHN), 4.47-4.70 (1H, m, CHN), 8.14 (1H, bs, NH), 8.22 (1H, bs, NH).Analysis calculated for C₁₀ H₁₈ N₂ S₂ : C, 52.17; H, 7.83; N, 12.17.Found C, 51.93; H, 7.44; N, 12.09.

EXAMPLE 22 1 H-Thieno[3,4-d]imidazole-hexahydro-2-thioxo-4-pentyl; 3aS(3aα,4β,6aα)

3H,5H-imidazo[1,5c]thiazole,tetrahydro-3,3-pentamethylene-7-hydroxymethyl-5-thioxo-1-pentyl 1S(1α,7β,7aα) (580 mg, 1.77 mmol) was dissolved in trifluoroacetic acid(4.6 ml) and water (1.2 ml) and the resultant solution was heated at100°-105° C. for 4 hours. The reaction mixture was cooled to 50° C. andconcentrated in vacuo. Ethanol (6.0 ml) was added and the solution wasagain concentrated under reduced pressure. The white residue wastriturated with ethyl acetate to afford 267 mg (66%) of1H-thieno[3,4-d]imidazole-hexahydro-2-thioxo-4-pentyl, 3aS (3aα,4β,6aα),mp 262°-262.5α_(D) ²⁵° C. =133° (C=0.01, TFA); IR (KBr) 3220, 2919; NMR(d, DMSO) 0.68-1.04 (3H, m, CH₂ CH₃), 1.08-1.96 (8H, m, C-CH₂),2.64-2.98 (2H, m, CH₂ S), 3.06-3.36 (1H, m, CHS), 4.28-4.46 (1H, m,CHN), 4.47-4.70 (1H, m, CHN), 8.14 (1H, bs, NH), 8.22 (1H, bs, NH).

EXAMPLE 23 1-H-Thieno[3,4-d]imidazole-hexahydro-2-oxo-4-pentyl;(3aα,4β,6aα)

To an ethanol solution (14 ml) containing1-H-thieno[3,4d]imidazole-hexahydro-2-thioxo-4-pentyl (3aα,4β,6aα) (307mg, 1.33 mmol) was added bromoethanol (208 ml, 2.93 mol). The solutionwas heated under reflux for 20 hours. Aqueous saturated sodium carbonate(1.5 ml) was added and the reaction was heated for an additional 10minutes, cooled, and concentrated in vacuo to give a solid residue whichwas dissolved in ethyl acetate (100 ml) and shaken with brine (2×35 ml).The organic extract was dried over magnesium sulfate, filtered andconcentrated in vacuo to afford white solids which were recrystallizedfrom ethyl acetate to give 196 mg (69%) of1-H-thieno[3,4-d]imidazole-hexahydro-2-oxo-4-pentyl (3aα,4β,6aα) mp144°-145° C. IR(KBr) 3175, 2899, 1709; NMR (d, DMSO) 0.69-1.10 (3H, m,CH₃), 1.11-1.86 (8H, m, CH₂), 2.44-2.96 (2H, m, CH₂ --S), 2.98-3.24 (1H,m, CHS), 4.02-4.21 (1H, m, CHN), 4.30-4.44 (1H, m, CHN), 6.36 (1H, bs,NH), 6.58 (1H, bs, NH); Mass spectrum: Calculated 214.1140, Observed:214.1144. Analysis Calculated for C₁₀ H₁₈ N₂ OS: C, 56.08; H, 8.41; N,13.08. Found C, 56.00; H, 8.07; N, 12.71.

EXAMPLE 24 (dl-Biotin)

A diglyme solution (3 ml) containing dl-thioxobiotin (255 mg, 0.981 mol)and bromoethanol (140 μl, 1.98 mmol) was allowed to reflux (150°) for2.5 hours. The solution was cooled, dilute sodium carbonate (50 ml) wasadded, and the solution was extracted with hexane (50 ml). The pH of theaqueous phase was adjusted to 1.5 with 6N HCL and extracted with ethylacetate (4×100 ml). The ethyl acetate extract was dried over magnesiumsulfate, filtered and concentrated in vacuo to give 148 mg (62%) ofcrude biotin, mp 220°-223° C. which was recrystallized from water IR(KBr) 3279, 2899, 1724; NMR (d, DMSO) 0.76-1.95 (6H, m, C--CH₂),2.00-2.40 (2H, t, CH₂ --CO), 2.70-2.98 (1H, m, CH₂ S) 3.00-3.54 (2H, m,CH₂ S, CHS) 4.00-4.44 (2H, m, CHN), 6.44 (1H, bs, NH), 6.55 (1H, bs,NH), 11.8-12.4 (1H, bs, OH). Analysis calculated for C₁₀ H₁₆ O₃ N₂ S: C,49.18; H, 6.56; N, 11.48; Found: C, 49.30; H, 6.34; N, 11.37. Massspectrum: calculated: 244.0880, found: 244.0925.

EXAMPLE 25 2,2-Dimethyl-5-pentyl-3-thiazoline

The procedure of M. Thiel, F. Asinger, K. Schmiedel, (Liebigs Ann. Chem.611, 121 (1958)) was employed. To a methanol solution (3 liters)containing sodium methoxide (216.7 g, 4.01 mole) at -10° C. was addedhydrogen sulfide (˜1 lb) until the solution was saturated. To thissolution was added dropwise over a 2 hour period 2-bromoheptaldehyde(775 g, 4.01 mol). The temperature of the reaction was maintained atabout -10° C. after the addition, acetone (734 g, 12.6 mole) was addedover a 10 minute period and the solution was stirred for an additional20 minutes at about -10° C. at which time ammonia was added over aperiod of 1.5 hours. The solution was then poured into water (4 liters)and extracted with ether (4×1 liter). The combined etheral extracts werewashed with brine (1×1 liter), dried over magnesium suflate andconcentrated in vacuo to afford after distillation (70°-80°/1.5 mm) 549g (74%) of 2,2-dimethyl-5-pentyl-3-thiazoline; NMR (d, CDCl₃) 0.6-2.2(11H, m, CH₂,CH₃) 1.70 (6H, s, C(CH₃)₂), 4.3-4.65 (1H, m, CHS), 7.10(1H, d, CHN).

EXAMPLE 26 3-Thiazoline-5-pentanoic acid, 2,2-dimethyl, methyl ester

The procedure of M. Thiel, F. Asinger, K. Schmiedel, (Liebigs Ann. Chem.611, 121 (1958)) was employed. A sodium methoxide solution was preparedby adding sodium (736 mg, 32 mmol) to methanol (30 ml) under a nitrogenatmosphere. This solution was cooled to -10° C. and saturated withhydrogen sulfide (˜20 minutes). To this solution was added dropwise overa 15 minute period 5-bromo-6-formylhexanoic acid, methyl ester (7.49 g,32 mmol). The reaction temperature was kept below -10° C. This solutionwas stirred for an additional 5 minutes at -10° C. at which time acetone(8 ml), was added. The reaction was stirred at -10° for 10 minutes atwhich time ammonia was introduced. The reaction temperature was notallowed to exceed 25° C. The reaction mixture became clear and ammoniawas bubbled into the solution for about 40 minutes. Water (100 ml) wasthen added and this solution was extracted with ether (4×100 ml). Theorganic layer was dried over magnesium sulfate, filtered, andconcentrated in vacuo to afford, after distillation (116°-125°/0.25 mm)6.19 g (85%) of 3-thiazoline-5-pentanoic acid, 2,2-dimethyl, methylester. IR (CHCL₃) 2944, 1734 1648. NMR (d, CDCL₃) 1.2-2.1 (12H, m,C--CH₂, C(CH₃)₂), 2.2-2.5 (2H, m, CH₂ CO), 3.7 (3H, s, OCH₃), 4.2-4.7(1H, m, CHS), 7.0-7.2 (1H, m, CHN).

EXAMPLE 27 2,2-pentamethylene-5-pentyl-3-thiazoline

The procedure of M. Thiel, F. Asinger, K. Schmiedel, (Liebigs Ann. Chem.611, 121 (1958)) was employed. To a methanol solution (75 ml) containingsodium hydrogen sulfide-water (6.25 g, 67.9 mmol) at -10° to 15° C. wasadded over a 15 minute period a methanol solution (15 ml) containing2-bromoheptaldehyde (13.1 g, 67.9 mol). The temperation of the reactionmixture was maintained at about -10° C. After stirring for 15 minutes,cyclohexanone (21.1 ml, 204 mmol) was added over a period of 2 minutes.This solution was stirred for an additional 15 minutes at -10° C. atwhich time ammonia was introduced. Ammonia was added over a one hourperiod and the reaction mixture was allowed to warm to room temperature.The clear solution was poured into water (250 ml) and extracted withether (3×200 ml). The organic extract was dried over magnesium sulfate,concentrated in vacuo and distilled (115°-125°/0.15 mm) to afford 10 g(66%) of 2,2-pentamethylene-5-pentyl-3-thiazoline. IR (CHCl₃) 2900,2830, 1648, 1530; NMR (d, CDCL₃) 0.68-2.9 (21H, m, CH₂, CH₃), 4.15-4.50(1H, m, CHS), 7.2 (1H, d, CHN).

EXAMPLE 28 1H-Thieno[3,4-d]imidazole-hexahydro-2-oxo, (3aα,4β,6aα)

To a methanol solution (20 ml) containing3H,5H-imidazo[1,5c]thiazole-7-carboxylic acid,tetrahydro-3,3-dimethyl-5-oxo-1-pentyl, ethyl ester (1α,7β,7aα) (358 mg,1.14 mol) was added potassium hydroxide (75 mg, 1.14 mmol) in water (20ml). The solution was stirred for 3 hours at room temperature,concentrated in vacuo, dissolved in ethyl acetate (200 ml) and extractedwith 6N HCL (50 ml). The organic extract was dried over magnesiumsulfate, filtered and concentrated under reduced pressure to give 330 mgof crude 3H,5H-imidazo[1,5c]thiazole. 7-carboxylic acid,tetrahydro-3,3-dimethyl-5-oxo-1-pentyl (1α,7β,7aα). IR (KBr), 3359,2927, 1733; NMR (d, DMSO) 0.7-2.3 (17H, m, CH₂, CH₃), 3.0-4.7 (3H, m,CHN, CHS), 6.7-7.03 (1H, m, NH). This acid was dissolved intrifluoroacetic acid (10 ml), deuterium oxide (0.6 ml) was added and thesolution was heated at 45° C. for 15 hours. The reaction mixture wasconcentrated in vacuo, taken up in ethyl acetate (200 ml) washed withwater (2×50 ml), dried over magnesium sulfate, filtered, concentrated invacuo, and precipitated with 1:1 methylene chloride:ether to afford 125mg of crude 1H-thieno[3,4-d]imidazole-2,4-dione, tetrahydro-6-pentyl-,(3aα,6β,6aα). An analytical sample, mp 247-247.5 was obtained after ahexane:methylene chloride recrystallization. IR (KBr) 3333, 2899, 1695.NMR (d, CDCL₃) 0.62-1.02 (3H, m, CH₃), 1.11-2.02 (8H, m, CH₂), 3.92-4.53(3H, m, CH), 6.66-6.90 (1H, m, NH), 7.30-7.56 (1H, m, NH). AnalysisCalculated for C₁₀ H₁₆ N₂ O₂ S: C, 52.63; H, 7.02; N, 12.28. Found: C,52.30; H, 7.00; N, 12.28. This lactone (174 mg, 0.76 mmol) was dissolvedin methanol (15 ml) at 0° C. Sodium borohydride (114 mg, 3.05 mmol) wasadded and the reaction mixture was allowed to warm to room temperature,stirred for an additional 1 hour and was then concentrated in vacuo. Theresidue was taken up in ethyl acetate (50 ml) and washed with water(1×20 ml), brine (1×20 ml), dried over magnesium sulfate, filtered andconcentrated to afford 70 mg of a white solid which was dissolved inacetic acid (15 ml) and treated with zinc (excess) at room temperature(4 hours), 40° C. (2 hours) and at reflux (3 hours). Thin layerchromatography (90:10:1 chloroform:methanol:ammonium hydroxide)indicated the formation of1-H-thieno[3,4-d]imidazole-hexahydro-2-oxo-4-pentyl (3aα,4β,6aα).

EXAMPLE 29 1H-thieno[3,4-d]imidazole-2,4-dione, tetrahydro-6-pentyl(3aα,6β,6aα)

To a methanol solution (20 ml) at 5° C. containing3H5H-imidazo[1,5c]thiazole-7-carboxylic acid,tetrahydro-3,3-dimethyl-5-oxo-1-pentyl, ethyl ester (1α,7α,7aα) (629 mg,2.0 mmol) was added potassium hydroxide (129 mg, 2.0 mmol) in water (2ml). The reaction mixture was stirred for 1 hour at room temperature,acidified to pH 3 with 1N HCL, extracted with ethyl acetate (3×100 ml),dried over magnesium sulfate, filtered and concentrated in vacuo toafford a white solid which was triturated with hexane:ether to give 327mg of crude 3H,5H-imidazo[1,5c]thiazole-7-carboxylic acid,tetrahydro-3,3-dimethyl-5-oxo-1-pentyl (1α,7α,7aα) mp 210°-211° C. IR(KBr) 3359, 2927, 1733; NMR (DMSO) 0.7-2.3 (17H, m, CH₂,CH₃), 3.0-4.7(3H, m, CH), 6.7-7.03 (1H, m, NH). Analysis Calculated for C₁₃ H₂₂ N₂ O₃S: C, 54.52; H, 7.74; N, 9.78. Found C, 54.50; H, 7.67; N, 10.05. Thisacid was dissolved in trifluoroacetic acid (4 ml), deuterium oxide (1ml) was added and the solution was stirred at room temperature for 2hours. The reaction mixture was concentrated in vacuo (3×) (toluene wasused to azeotrope off water) to afford 289 mg of crude thiol which wasplaced in dry methylene chloride (125 ml) in the presence oftriethylamine (170 ml, 1.22 mol) at 0° C. Ethylchloroformate (117 ml,1.22 mmol) was added and the reaction mixture was stirred for 2 hours atroom temperature. The solvent was then concentrated under reducedpressure, and the white residue was taken up in ethyl acetate (80 ml)).The organic solution was washed with water (40 ml), dried over magnesiumsulfate, filtered, and concentrated in vacuo to afford 270 mg of solidswhich were recrystallized from ethyl acetate to afford 110 mg of1H-thieno[3,4-d]imidazole-2,4-dione, tetrahydro-6-pentyl (3aα,6α,6aβ),mp 177°-180° C. IR (KBr) 3125, 2890, 1786, 1670.

Analysis Calculated for C₁₀ H₁₆ N₂ O₂ S: 52.63; H, 7.02; N, 12.28.Found: 52.88; H, 7.15; N, 12.22. This thiolactone (21 mg, 0.09 mmol) wasdissolved in tetrahydrofuran (2 ml), DBU 1,8-diazabicyclo[5.4.0]undecC-7 ene (1.38 μl, 0.009 mmol) was added and the solution was stirred for20 minutes. IN HCL (100 l, 0.1 mol) was added and the reaction mixturewas concentrated under reduced pressure, taken up in ethyl acetone (50ml) and extracted with water. The organic layer was dried over magnesiumsulfate, filtered and concentrated in vacuo to afford 21 mg of1H-thieno[3,4d]imidazole-2,4-dione, tetrahydro-6-pentyl-(3aα,6β,6aα).

I claim:
 1. A method for preparation of biotin comprising(a) contactingin solution a compound of the formula ##STR15## wherein X is sulfur oroxygen;R₁ is --(CH₂)₄ CH₃, or --(CH₂)₃ OR or --(CH₂)₅ OR wherein R isalkyl, or --(CH₂)₄ CN, or --(CH₂)₄ COOR' wherein R' is alkyl or phenyl;R₂ and R₃ when taken together, are cycloalkyl or --CH₂ --CH₂ --Y--CH₂--CH₂ wherein Y is sulfur, oxygen or NR" wherein R" is COOR'" whereinR'" is alkyl, or R₂ and R₃ when taken separately, are each alkyl,cycloalkyl or phenyl, provided that R₂ and R₃ are not both phenyl; andR₄ is alkyl, alkoxyalkyl, cycloalkyl, monoalkyl substituted cycloalkyl,phenyl or mono-, di or trialkyl substituted phenyl; said alkyl andalkoxy having from 1 to 4 carbon atoms and said cycloalkyl having from 5to 7 carbon atoms, with an alkali metal borohydride in a protic ornon-protic solvent followed by addition of water; (b) contacting theresultant hydroxy compound with strong aqueous acid or contacting theresultant hydroxy compound in non-protic solvent with an alkyl or arylsulfonyl halide or with an acyl halide in the presence of a weak baseand contacting the product with strong aqueous acid; (c) refluxing whenX is sulfur, the resultant bicyclic thiourea with a halo alcohol havingfrom 2 to 4 carbon atoms; and (d) when R₁ is --(CH₂)₄ CH₃, oxidizing theresultant product or, when R₁ is --(CH₂)₄ COOR', hydrolyzing theresultant product, when R₁ is --(CH₂)₄ CN, hydrolyzing the resultantproduct, or when R₁ is --(CH₂)₅ OR, hydrolyzing and oxidizing theresultant product, or when R₁ is --(CH₂)₃ OR(1) treating with aceticacid saturated with hydrogen bromide followed by sodium diethylmalonate, (2) hydrolyzing the resultant diester with barium hydroxide,and (3) heating at a temperature of about 180° until reaction issubstantially complete.
 2. The method of claim 1 wherein said metalborohydride is sodium borohydride.
 3. The method of claim 1 wherein saidweak base is trimethylamine, triethylamine or pyridine.
 4. The method ofclaim 1 wherein said acid is trifluoroacetic acid or methanesulfonicacid.
 5. The method of claim 1 wherein said halo alcohol isbromoethanol.
 6. The method of claim 1 wherein said borohydridederivative is sodium borohydride, said sulfonyl chloride ismethanesulfonyl chloride or camphorsulfonyl chloride, said acyl halideis acetyl chloride, said weak base is triethylamine, and said acid istrifluoroacetic acid.
 7. The method of claim 1 whereinR₁ is --(CH₂)₄ CH₃or --(CH₂)₄ COOR' wherein R' is alkyl; R₂ and R₃ when taken together arecycloalkyl, or R₂ and R₃ when taken separately are each alkyl; and R₄ isalkyl.
 8. The method of claim 7 whereinR₁ is --(CH₂)₄ CH₃ or --(CH₂)₄COOCH₃ ; R₂ and R₃ when taken together are cyclohexyl, or R₂ and R₃ whentaken separately are each methyl; and R₄ is ethyl.
 9. The method ofclaim 8 wherein R₁ is --(CH₂)₄ CH₃, R₂ and R₃ when taken together arecyclohexyl, X is sulfur and R₄ is ethyl.
 10. A method for preparation ofd-biotin comprising(a) contacting a compound of the formula ##STR16##wherein X is sulfur or oxygen; R₁ is --(CH₂)₄ CH₃, or --(CH₂)₃ OR or--(CH₂)₅ OR wherein R is alkyl, or --(CH₂)₄ CN, or --(CH₂)₄ COOR'wherein R' is alkyl or phenyl;R₂ and R₃ when taken together arecycloalkyl or --CH₂ --CH₂ --Y--CH₂ --CH₂ wherein Y is sulfur, oxygen orNR" wherein R" is COOR'" wherein R'" is alkyl, or R₂ and R₃ when takenseparately are each alkyl, cycloalkyl or phenyl, provided that R₂ and R₃are not both phenyl; said alkyl having from 1 to 4 carbon atoms and saidcycloalkyl having from 5 to 7 carbon atoms; with (d) or (l)camphorsulfonyl chloride in non-protic solvent in the presence of weakbase; (b) separating the resultant diastereomeric mixture ofthiazolidine O-camphorsulfonates to provide a compound of the formula##STR17## wherein R₅ is camphorsulfonyl, (c) hydrolyzing thethiazolidine moiety of the requisite O-camphorsulfonyl stereoisomer inthe presence of acid; (d) when X is sulfur, treating the resultantbicyclic thiourea with a haloalcohol having from 2 to 4 carbon atoms;and (e) when R₁ is --(CH₂)₄ CH₃, oxidizing the resultant product or,when R₁ is --(CH₂)₄ COOR', hydrolyzing the resultant product or when R₁is --(CH₂)₄ CN, hydrolyzing the resultant product or, when R₁ is--(CH₂)₅ OR, hydrolyzing and oxidizing the resultant product, or when R₁is --(CH₂)₃ OR(1) treating with acetic acid saturated with hydrogenbromide followed by sodium diethyl malonate (2) hydrolyzing theresultant diester with barium hydroxide, and (3) heating at atemperature between 150° to 200° until reaction is substantiallycomplete.
 11. The method of claim 10 wherein said diastereomeric mixtureis separated by contacting with silica gel or alumina.
 12. The method ofclaim 10 wherein R₁ is --(CH₂)₄ CH₃, X is sulfur and R₂ and R₃ togetherform cyclohexyl.